Cancer cells exhibit profound metabolic differences from their normal counterpart. Currently, there is significant optimism that the metabolic traits unique to cancer cells may represent viable therapeutic targets. Work from a number of groups, ours included, has firmly implicated the MYC oncoprotein as a central regulator of the metabolic reprogramming that takes place in human cancer cells. To gain insight into MYC's role in human cancer, we recently performed an unbiased expression profiling screen to identify essential downstream targets whose transcription is tightly correlated with malignant transformation. Via the screen, we identified the POLRMT gene. POLRMT is a nuclear gene that encodes the single chain RNA polymerase molecule, mtRNAP that is responsible for transcription from the small, circular mitochondrial genome (mtDNA). (MYC had previously been shown to regulate the three major nuclear RNA polymerase molecules.) Functional studies of the induction of POLRMT/mtRNAP by MYC show that MYC regulates all transcription within the mitochondria via its control of POLRMT expression. Furthermore, because mtRNAP transcribes the RNA primers required for replication of the mtDNA, MYC regulates cellular mitochondrial genome content as well. Genetic rescue experiments show that both of these efects depends exclusively on mtRNAP and not on a recently described nuclear isoform encoded by POLRMT, spRNAP-IV. The link between MYC and POLRMT expression has been confirmed in a variety of human cancer cell lines and is supported by evidence from public cancer gene expression datasets. MYC also regulates POLRMT expression in primary human cells. Most importantly, blocking the ability of MYC to induce POLRMT expression results in robust apoptosis that strictly depends on the presence of activated MYC. Strategies that reactivate the latent apoptotic potential or "intrinsic tumor suppressor activity" of MYC are widely viewed as an exciting opportunity for future anti-cancer therapy. Finally, the MYC-dependent apoptosis that results from blocking POLRMT induction appears to result from a unique requirement for POLRMT/mtRNAP in the expression of mitochondrial-encoded components of the electron transport chain and subsequent effects on the pyrimidine biosynthesis pathway. Targeting pyrimidine biosynthesis has been recognized as a successful anti-cancer therapy for many decades and our data suggest that induction of POLRMT/mtRNAP may represent a novel node for intervention in this pathway, specifically in the forms of human cancer that express high levels of MYC. The studies proposed here will test this hypothesis and provide a detailed mechanistic understanding of the pathway we have identified, along with the essential, MYC-dependent survival function it provides to tumor cells. PUBLIC HEALTH RELEVANCE: Cancer cells have a unique set of metabolic reactions relative to their normal counterparts and many current efforts are focused on exploiting these differences for anti-cancer therapy. We have identified a unique mitochondrial enzyme regulated by the MYC oncoprotein that is essential for survival of cancer cells and which represents a targetable node in the pathway MYC controls.